Antiparastic auermectin derivatives

ABSTRACT

Antiparasitic avermectin derivatives of formula (I), where the broken line represents an optional bond, R 1  and R 4  are independently H, OH, halo, oximino, or an organic radical, R 2 , R 6  and R 7  are organic radicals and R 3  is alpha-oleandrosyl or 4&#39;-(alpha-oleandrosyl)-alpha-oleandrosyl optionally substituted at the 4&#39;- or 4&#34;-position, and R 12  and R 13  are independently H, CN, CONH 2 , C 1  -C 8  alkyl or aryl optionally substituted with at least one halo, OH, C 1  -C 8  alkylthio group.

This application is a continuation of application Ser. No. 08/591,534,filed on Jan. 25, 1996, now adandoned entitled "Antiparasitic Agents",-which is the national stage of PCT Application No. EP 94/02433, filedon Jul. 22, 1994.

This invention relates to antiparasitic agents and in particular tocompounds related to the avermectins and milbemycins but having analkylidene substituent at the 5-position.

The avermectins are a group of broad-spectrum antiparasitic agentsreferred to previously as the C-076 compounds They are produced byfermenting a certain strain of micro-organism Streptomyces avermitilisin an aqueous nutrient medium. The preparation and structure of thesecompounds obtained by fermentation are described in British PatentSpecification 1573955. The milbemycins are structurally relatedmacrolide antibiotics lacking the sugar residues at the 13-position.They may be produced by fermentation, for example as described inBritish Patent Specification No. 1390336 and European PatentSpecification No. 0170006.

Compounds related to the original C-076 avermectins have also beenprepared by fermentation of avermectin-producing micro-organisms. Forexample European Patent Specifications 0214731 and 0317148 describeproduction of compounds related to the C-076 avermectins but having adifferent substituent at the 25-position by fermentation in thepresence, in the fermentation medium, of certain acids.

In addition to these fermentation-derived products, a large number ofpublications describe compounds derived semisynthetically from theseproducts, many of which possess useful antiparasitic properties. Some ofthis chemistry is reviewed in Macrolide Antibiotics, Omura S., Ed.,Academic Press, New York (1984) and by Davies, H. G. and Green, R. H. inNatural Product Reports (1986), 3, 87-121 and in Chem Soc Rev (1991),20, 211-269 and 271-239.

Other publications mentioning different combinations of substituents atvarious positions on the avermectin or milbemycin nucleus areEP-A-317148, 340932, 355541, 350187, 410165, 259779 and 254583;DE-A-2329486 and GB-A-2166436.

No avermectin derivatives having an alkylidene substituent at the5-position are known, neither has any process capable of producing suchcompounds been reported. Japanese Patent Application No. 86-94754(published under No. 87-252788) of Sankyo describes milbemycins andaglycone derivatives having a methylidene substituent at the 5-position,but the processes described for making them cannot be used for makingsimilarly substituted avermectins or avermectin monosaccharides, assaccharide groups are removed by hydrolysis under the acidic conditionsused.

It has now been discovered that avermectin derivatives and theirmonosaccharides having an alkylidene substituent at the 5-position maybe prepared and that certain of these compounds have unexpectedantiparasitic properties, in particular high potency against importantarthropod parasites of cats and dogs. In addition, they have improvedsafety in mammals compared to previously known avermectins.

According to one aspect of the invention, there are provided compoundsof formula (I) having antiparasitic activity ##STR1## wherein the brokenline represents an optional bond, R¹ and R⁴ being absent when this bondis present, R¹ and R⁴ are independently H or OR¹⁴ where R¹⁴ is H, C₁ -C₈alkyl, C₂ -C₈ alkenyl, aralkyl, C₂ -C₈ alkanoyl, C₃ -C₈ alkenoyl,aralkanoyl, aroyl or carbamoyl;

R² is:

(a) an alpha-branched C₃ -C₈ alkyl, alkenyl, alkoxy-alkyl, oralkylthioalkyl group; and alpha-branched C₄ -C₈ alkynyl group; a (C₅-C₈)cycloalkyl-alkyl group wherein the alkyl group is an alpha-branchedC₂ -C₅ alkyl group; a C₃ -C₈ cycloalkyl or C₅ -C₈ cycloalkenyl group,either of which may optionally be substituted by methylene or one ormore C₁ -C₄ alkyl groups or halo atoms; or a 3 to 6 membered oxygen orsulphur containing heterocyclic ring which may be saturated, or fully orpartially unsaturated and which may optionally be substituted by one ormore C₁ -C₄ alkyl groups or halo atoms; or

(b) a group of the formula --CH₂ R⁸ wherein R⁸ is H, C₁ -C₈ alkyl, C₂-C₈ alkenyl, C₂ -C₈ alkynyl, alkoxyalkyl or alkylthioalkyl containingfrom 1 to 6 carbon atoms in each alkyl or alkoxy group, wherein any ofsaid alkyl, alkoxy, alkenyl or alkynyl groups may be substituted by oneor more halo atoms; or R⁸ is a C₃ -C₈ cycloalkyl or C₅ -C₈ cyoloalkenylgroup, either of which may optionally be substituted by methylene or oneor more C₁ -C₄ alkyl groups or halo atoms; or R⁸ is a 3 to 6 memberedoxygen or sulphur containing heterocyclic ring which may be saturated,or fully or partially unsaturated and which may optionally besubstituted by one or more C₁ -C₄ alkyl groups or halo atoms; or R⁸ is agroup of the formula SR⁹ wherein R⁹ is C₁ -C₈ alkyl, C₂ -C₈ alkenyl, C₃-C₈ alkynyl, C₃ -C₈ cycloalkyl, C₅ -C₈ cycloalkenyl, phenyl orsubstituted phenyl wherein the substituent is C₁ -C₄ alkyl, C₁ -C₄alkoxy or halo; or R⁸ is a 3 to 6 membered oxygen or sulphur containingheterocyclic ring which may be saturated, or fully or partiallyunsaturated and which may optionally be substituted by one or more C₁-C₄ alkyl groups or halo atoms; or

(c) a C₁ -C₆ alkyl group substituted by one oxo or one or more hydroxygroups or by a single oxygen atom on two adjacent carbon atoms formingan oxirane ring, or R² is a C₁ -C₅ alkyl group substituted by a (C₁ -C₆)alkoxy-carbonyl group, said substituents on R₂ being attached to eitheror both of a terminal carbon atom and a carbon atom adjacent a terminalcarbon atom of R² ; or

(d) ═CH₂ or a group of the formula: ##STR2## wherein R¹⁰ and R¹¹ areboth H; R¹⁰ is H and R¹¹ is C₁ -C₃ alkyl, or one of R¹⁰ and R¹¹ is H andthe other is phenyl, heteroaryl, C₂ -C₆ alkoxycarbonyl or substitutedphenyl or heteroaryl wherein said substituent is fluorine, chlorine, C₁-C₄ alkyl, C₁ -C₄ alkoxy, C₁ -C₄ alkylthio, hydroxy(C₁ -C₄)alkyl, cyano,aminosulphonyl, C₂ -C₆ alkanoyl, C₂ -C₆ alkoxycarbonyl, nitro,trifluoromethyl, trifluoromethoxy, amino or mono or di(C₁ -C₄)alkylamino; and X is a direct bond or is an alkylene group having from 2to 6 carbon atoms which may be straight or branched-chain; or

(e) phenyl which may optionally be substituted with at least onesubstituent selected from C₁ -C₄ alkyl, C₁ -C₄ alkylthio groups, haloatoms, trifluoromethyl, and cyano;

or R² may be a group of formula (II): ##STR3## wherein Z is O, S or--CH₂ -- and a, b, c and d may each independently be 0, 1 or 2, the sumof a, b, c and d not exceeding 5;

R⁶ is H or C₁ -C₆ alkyl;

R⁷ is CH₃, --CH₂ --OR¹⁴ where R¹⁴ is as defined above, or --CH₂ X whereX is a halogen atom; and

R³ is a group of formula: ##STR4## wherein R⁵ is attached to C-4" orC-4' by a single bond and is hydrogen, halo, hydroxy, C₁ -C₉ alkanoyloxyor alkenoyloxy, aroyloxy, C₁ -C₈ alkoxy, amino, N-(C₁ -C₈)alkylamino,N,N-di(C₁ -C₉)-alkylamino, N-(C₁ -C₉)alkanoylamino, or N,N-di(C₁-C₉)alkanoylamino;

or R⁵ is attached to C-4" or C-4' by a double bond and is oxo,optionally substituted oximino, semicarbazido, N-(C₁-C₄)alkylsemicarbazono, N,N-di(C₁ -C₄)alkylsemicarbazono, (C₁-C₄)alkylbenzoylhydrazono, and R¹² and R¹³ are independently H, CN,CONH₂, C₁ -C₆ alkyl or aryl optionally substituted with at least onehalo, OH, C₁ -C₆ alkoxy or C₁ -C₆ alkylthio group.

In all the above definitions, unless the context requires otherwise,alkyl groups containing 3 or more carbon atoms may be straight orbranched-chain; halo means fluoro, chloro, bromo or iodo; and aryl meansphenyl optionally substituted by one or more C₁ -C₄ alkoxy groups orhalo atoms.

Compounds within the scope of the invention include

5-cyanomethylidene-25-cyclohexyl avermectin B2;

5-carbamoylmethylidene-25-cyclohexyl avermectin B2;

5-cyanomethylidene-22,23-dihydroavermectin B1a monosaccharide;

5-methylidene-22,23-dihydroavermectin B1a monosaccharide;

5-methylidene-25-cyclohexyl-22,23-dihydroavermectin B1 monosaccharide;

5-methylidene-25-cyclohexylavermectin B2;

and 5-ethylidene-25-cyclohexylavermectin B2.

The compounds of formula (I) may be prepared from a compound of formula(II): ##STR5## wherein R¹, R², R³, R⁴, R⁶ and R⁷ are as defined above.The compounds of formula (II) may generally be prepared by oxidation ofthe corresponding 5-hydroxy compound, for example using manganesedioxide, groups R¹ -R⁷ being protected by conventional methods ifnecessary. These 5-hydroxy analogues may themselves be prepared bymethods known in the art.

The addition of a phosphorus ylide of the formula R¹³ R¹² C=PPh₃ to acompound of the formula (II) at low temperature (-100° C. to 0° C.) inan inert organic solvent such as tetrahydrofuran produces a compound offormula (I). The phosphorus ylide may be prepared using known methodsfrom a compound of formula R¹³ R¹² CH⊕PPh₃ X⊖, where X⊖ is a halide ion,in the presence of base. Alternatively a compound of formula R¹³ R¹²CHP(O)(OR¹⁵)₂, where R¹⁵ is an alkyl group, may firstly be treated witha base, then added to a compound of formula (II), to produce a compoundof formula (I). This reaction may also be performed in a two-phasemixture of a chlorinated organic solvent and aqueous alkali containing acompound of formula R¹³ R¹² CHP(O)(OR¹⁵)₂ and a phase-transfer reagentand a compound of formula (II), to form a compound of formula (I).

When it is desired to prepare a compound in which R¹² or R¹³ iscarbamoyl, a phosphorus ylide in which R¹² or R¹³ is cyano may be usedto obtain a cyanomethylidene derivative of formula (I) which may behydrolysed under mild conditions, for example in the presence ofmanganese dioxide in methylene chloride at ambient temperature.

The compounds of the invention are effective in treating a variety ofconditions caused by endoparasites including, in particular,helminthiasis which is most frequently caused by a group of parasiticworms described as nematodes and which can cause severe economic lossesin swine, sheep, horses and cattle as well as affecting domestic animalsand poultry. The compounds are also effective against other nematodeswhich affect humans and various species of animals including, forexample, Dirofilaria in dogs and various parasites which can infectanimals and humans including gastro-intestinal parasites such asAncylostoma, Necator, Ascaris, Strongyloides, Trichinella, Toxocara,Capilaria, Trichuris, Enterobius and parasites which are found in theblood or other tissues and organs such as filiarial worms and theextra-intestinal stages of Strongyloldes, Trichinella and Toxocara.

The compounds are also of value in treating ectoparasite infectinsincluding in particular arthropod ectoparasites such as fleas, ticks,mites, lice, blowfly and biting Insects and migrating dipterous larvaewhich can affect cattle and horses.

The compounds are also insecticides active against household pests suchas the cockroach, clothes moth, carpet beetle and the housefly as wellas being useful against arthropod pests of stored grain and ofagricultural plants such as spider mites, aphids, caterpillars andagainst migratory orthopterans such as locusts. We have discovered thatcertain compounds within the scope of this invention have unexpectedlyhigh potent activity against important arthropod parasites of cats anddogs.

The compounds of formula (I) may be administered as a formulationappropriate to the specific use envisaged and to the particular speciesof host animal being treated and the parasite or insect involved. Foruse as an anthelmintic the compounds may be administered by injection,either subcutaneously or intramuscularly, alternatively they may beadministered orally in the form of a capsule, bolus, tablet, chewabletablet or liquid drench, or they may be administered as a topicalformulation or as an implant. For topical application dip, spray,powder, dust, pour-on, spot-on, jetting fluid, shampoos, collar, tag orharness may be used. Such formulations are prepared in a conventionalmanner in accordance with standard veterinary practice. Thus capsules,boluses or tablets may be prepared by mixing the active ingredient witha suitable finely divided diluent or carrier, additionally containing adisintegrating agent and/or binder such as starch, lactose, talc, ormagnesium stearate. A drench formulation may be prepared by dispersingthe active ingredient in an aqueous solution together with dispersing orwetting agents and injectable formulations may be prepared in the formof a sterile solution or emulsion. Pour-on or spot-on formulations maybe prepared by dissolving the active ingredient in an acceptable liquidcarrier vehicle, such as butyl digol, liquid paraffin or non-volatileester with or without addition of a volatile component such asisopropanol. Alternatively, pour-on, spot-on or spray formulations canbe prepared by encapsulation to leave a residue of active agent on thesurface of the animal. These formulations will vary with regard to theweight of active compound depending on the species of host animal to betreated, the severity and type of infection and the body weight of thehost. Generally for oral parenteral and pour-on administration a dose offrom about 0.001 to 10mg per g of animal body weight given as a singledose or in divided doses for a period of from 1 to 5 days will besatisfactory but of course there can be instances where higher or lowerdosage ranges are indicated and such are within the scope of thisinvention.

As an alternative the compounds may be administrated with the animalfeedstuff and for this purpose a concentrated feed additive or premixmay be prepared for mixing with the normal animal feed.

For use as an insecticide and for treating agricultural pests thecompounds are applied as sprays, dusts, pour-on formulations, emulsionsand the like in accordance with standard agricultural practice.

For human use the compounds are administered as a pharmaceuticallyacceptable formulation in accordance with normal medical practice.

The invention is illustrated by the following Examples, in which"avermectin B2" refers to an avermectin having an OH substituent at the23-position and a single bond at the 22-23 position, "avermectin B1"refers to an avermectin having a double bond at the 22-23 position,"avermectin B1a" is as for avermectin B1 and having a sec-butylsubstituent at the 25-position. An avermectin monosaccharide is anavermectin derivative having an alpha-oleandrosyl substituent at the13-position, the avermectins themselves having a 4¹-(alpha-oleandrosyl)-alpha-oleandrosyl group at this position.

NMR spectra were measured using a Bruker 300 MHz spectrometer. Massspectra were measured on a VG Mark I 7070E mass spectrometer using asample matrix of triethylene glycol with solid sodium chloride.

EXAMPLE 1 5-Cyanomethylidene-25-cyclohexylavermectin B2

Tetra-n-butylammonium bromide (0.5 g), dimethyl-cyanomethylphosphonate(3.5 ml), 1,3-dimethyl-3,4,5,6-tetrahydro-2-(1H)-pyrimidinone (1 ml),methylene chloride (50 ml), and a solution of sodium hydroxide (17 g) inwater (50 ml) were stirred together rapidly at 20° C. for 0.5 hours.Then the dichloromethane layer was separated and filtered throughphase-separating paper into a flask containing the compound ofPreparation 1 (0.5 g). The resultant solution was stirred at 20° C. for0.5 hour. The mixture was evaporated in vacuo at 40° C. and thendissolved in methylene chloride (50 ml) and then washed with saturatedammonium chloride solution (50 ml). The organic layer was separated anddried over sodium sulphate, then filtered and evaporated in vacuo. Theresidue was purified by chromatography on silica gel (50 g). Elution waswith a mixture of methylene chloride and methanol. The proportion ofmethanol was increased from 0 to 7% over 2 liters. 50 ml fractions werecollected and the product identified by thin-layer chromatography.(Rf=0.3 using a solvent system of 5% methanol in methylene chloride).Final purification was achieved by reverse-phase HPLC, using a C18Zorbax (Trademark, Dupont) column (21 mm×25 cm) eluting with a mixtureof methanol and water (80:20) at a flowrate of 9 mls per minute. 4.5 mlfractions were collected and fractions 118 to 130 were combined andevaporated to yield a compound of formula (I), wherein R¹ is OH, R² iscyclohexyl, R³ is 4'-(alpha-L-oleandrosyl)-L-oleandrosyloxy, R⁴ is H,the 22,23 double-bond is absent, R⁶ and R⁷ are methyl, R¹² is CN and R¹³is H. The compound has characteristic mass and NMR spectra:

Mass spectrum (FAB): 976 (MK+). NMR spectrum (300 MHz): δ (CDCl₃) ppm:6.1 (s,1H)} H-3 and H-5a. 6.12 (s,1H)}; 1.6 (s,3H, H-4a).

EXAMPLE 2 5-(Carbamoylmethylidene-25-cyclohexyl)avermectin B2

The product of Example 1 (250mg) was dissolved in methylene chloride(100 ml) and manganese dioxide (1.15 g) was added. The resultingsuspension was stirred at 20° C. for 1 day. Then a further 1 g ofmanganese dioxide was added and stirring continued for 2 days. Thenanother 1 g of manganese dioxide was added to the reaction mixture andstirring continued for 1 week. The mixture was then filtered through apad of celite and the filtrate evaporated. The residue was purified byreverse-phase HPLC, using a C18 Zorbax (Trademark, Dupont) column (21mm×25 cm) eluting with a mixture of methanol and water (75:25) at aflowrate of 9mls/min. The relevant fractions were combined to yield acompound of formula (I), wherein R¹ is OH, R² is cyclohexyl, R³ is4'-(alpha-L-olendrosyl)-L-oleandrosyloxy, R⁴ is H, the 22,23 double-bondis absent, R⁶ and R⁷ are methyl, R¹² is CONH₂ and R¹³ is H. The compoundhas characteristic mass and NMR spectra:

Mass spectrum (FAB): 994 (MK+). NMR spectrum (300 MHz): δ (CDCl₃) ppm:6.6 (s,1H)} H-3 and H-5a 6.2 (s,1H)}; 1.6 (s,3H, H-4a). 6.2 (br.s. 1H)}NH₂ of; 5.5 (br.s. 1H)} carbamoyl;

EXAMPLE 3 5-Cyanomethylidene-22,23-dihydroavermectin B1a monosaccharide

By the method of Example 1 the title compound was prepared from thecompound of Preparation 2. Purification was achieved by columnchromatography on silica gel (30 g) eluting with a mixture of ethylacetate in methylene chloride. The proportion of ethyl acetate wasincreased from 0 to 40% over 2 liters. Final purification was achievedby reverse-phase HPLC, using a C-18 Zorbax (Trademark, Dupont) column(21 mm×25 cm) eluting with a mixture of methanol and water (81:19) at aflowrate of 9 mls/min. 9 ml fractions were collected. Fractions 70 to 80were combined to yield a compound of formula (I), wherein R¹ is H, R² is2-butyl, R³ is L-oleandrosyloxy, R⁴ is H, the double bond is absent, R⁶and R⁷ are methyl, R¹² is CN and R¹³ is H. The compound hascharacteristic mass and NMR spectra:

Mass spectrum (FAB): 790 (MK+). NMR spectrum (300MHz): δ (CDCl₃) ppm:6.1 (s,1H)} H-3 and H-5a. 6.12 (s,1H)}; 1.55 (s,3H, H-4a).

EXAMPLE 4 5-Methylidene-22,23-dihydroavermectin B1a monosaccharide

To a stirred solution of methyl triphenylphosphonium bromide (430mg) indry tetrahydrofuran at 0° C. under nitrogen was added a solution ofbutyl lithium in hexanes (1.6M, 0.75ml ). The suspension was stirred for15 minutes at 0° C., then the compound of Preparation 2 (140 mg) in drytetrahydrofuran (10 ml) was added. The cooling bath was removed and thestirred mixture allowed to warm to 20° C. After stirring at thistemperature for 1 hour the reaction mixture was added to saturatedammonium chloride solution (50ml) and extracted with methylene chloride.The organic layer was separated and dried over sodium sulphate, thenfiltered and evaporated. The residue was purified by reverse-phase HPLC.The column used by a C18 Zorbax (Trademark, Dupont), (21 mm×25 cm),eluting with a mixture of methanol and water (86:14) at a flow rate of 9mls/min. 9ml fractions were taken and fractions 58 to 64 were combinedand evaporated to yield the compound of formula (I) wherein R¹ is H, R²is 2-butyl, R³ is L-oleandrosyloxy, R⁴ is H, the double bond is absent,R⁶ and R⁷ are methyl, R¹² and R¹³ are H. The compound has characteristicmass and NMR spectra:

Mass spectrum (FAB): 749 (MNa+). NMR spectrum (300MHz) δ (CDCl₃) ppm:5.55 (br.s., 2H, H-5a). 1.9 (br.s., 3H, H-4a).

EXAMPLE 5 5-Methylidene-25-cyclohexyl-22,23-dihydroavermectin B1monosaccharide

By the method of Example 4 the title compound was prepared from thecompound of Preparation 3. Purification was achieved by columnchromatography on silica gel (150 g); eluting with hexane, thenmethylene chloride, then ethyl acetate. Relevant fractions were combinedto yield, on evaporation, 760 mg of crude material. Further purificationwas achieved by reverse-phase HPLC on a Dynamax C-18 (Trademark, Rainin)column, (41.4 mm×25 cm) eluting with a mixture of methanol and water(88:12) at a flowrate of 40 mls/min. Fractions 29 to 34 were combinedand evaporated to yield 139 mg of a compound of formula (I) wherein R¹is H, R² is cyclohexyl, R³ is L-oleandrosyloxy, R⁴ is H, the double bondis absent, R⁶ and R⁷ are methyl, R¹² and R¹³ are H. The compound hascharacteristic mass and NMR spectra:

Mass spectrum (FAB): 775 (MNa+). NMR spectrum (300MHz): δ (CDCl₃) ppm:5.57 (br.s., 2H, H-5a). 1.95 (br.s., 3H, H-4a).

EXAMPLE 6 5-Methylidene-25-cyclohexylavermectin B2

By the method of Example 4, the title compound was prepared from thecompound of Preparation 1. Following silica gel chromatography, 80 mg ofa crude product was isolated. This was further purified by reverse-phaseHPLC on a Dynamax C-18 (Trademark, Rainin) column, (41.4 mm×25 cm)eluting with a mixture of methanol and water (85:15) at a flowrate of 40mls/min. 30 ml fractions were collected, and fractions 35 to 44 werecombined and evaporated to yield the compound of formula (I), (130 mg),wherein R¹ is OH, R² is cyclohexyl, R³ is4'-(alpha-L-oleandrosyl)-L-oleandrosyloxy, R⁴ is H, the 22,23 doublebond is absent, R⁶ and R⁷ are methyl and R¹² and R¹³ are H. The compoundhas characteristic mass and NMR spectra:

Mass spectrum (FAB): 930 (MNH₄ +). NMR spectrum (300 MHz): δ (CDCl₃)ppm: 5.55 (br.s., 2H, H-5a). 1.9 (br.s., 3H, H-4a).

EXAMPLE 7 5-Ethylidene-25-cyclohexylavermectin B2

The title compound was prepared by the method of Example 6, except thatethyl triphenylphosphonium bromide was used instead of methyltriphenylphosphonium bromide. The product was purified by reverse-phaseHPLC using a C18 Zorbax (Trademark, Dupont) column (21 mm×25 cm) elutingwith a mixture of methanol and water (84:16) at a flowrate of 9 mls/min.9 ml fractions were collected and fractions 86 to 90 were combined andevaporated to yield a compound of formula (I) wherein R¹ is OH, R² iscyclohexyl, R³ is 4'-(alpha-L-oleandrosyl)-L-oleandrosyloxy, R⁴ is H,the 22,23 double bond is absent, R⁶ and R⁷ are methyl, R¹² is H and R¹³is methyl. The compound has characteristic mass and NMR spectra:

Mass spectrum (FAB): 949 (MNa+). NMR spectrum (300 MHz): δ (CDCl₃) ppm:5.55 (br.s., 1H, H-5a). 2.15 (br.s., 3H, H-5b). 2.0 (br.s., 3H, H-4a).

PREPARATION 1 5-Keto25cyclohexylavermectin B2

To a suspension of manganese dioxide (3 g) in diethyl ether (50 ml) wasadded 25cyclohexylavermectin B2 (2 g), obtained as described inEP-A-214731. The mixture was stirred for 1 day, and a further portion ofmanganese dioxide (3 g) was added. After stirring for a second day at20° C. a third portion of manganese dioxide was added and the mixturestirred for a third day. The manganese dioxide was then removed byfiltration through Celite and the filtrate was evaporated to give 0.6 gof the title compound which has characteristic mass and NMR spectra:

Mass spectrum (FAB): 937 (MNa+). NMR spectrum (300 MHz): δ (CDCl₃) ppm:6.6 (br.s., 1H, H-3). 1.9 (br.s., 3H, H4a).

PREPARATION 2 5-Keto-22,23-dihydroavermectin B1a monosaccharide

By the method of Preparation 1, the title compound was prepared from25-(2-butyl)-22,23-dihydroavermectin B1a monosaccharide obtained asdescribed in U.S. Pat. No.-4,199,569.

PREPARATION 3 5-Keto-25cyclohexyl-22,23-dihydroavermectin B1monosaccharide

25-Cyclohexylavermectin B1 (9.9 g) was dissolved in toluene (1 liter)and Wilkinson's catalyst (tristriphenylphosphine rhodium (I) chloride)(9.25 g) was added. The solution was hydrogenated on a large Parr shakerat room temperature at 50 psi hydrogen pressure. After 3 hours thereaction vessel was depressurised and allowed to stand for 12 hoursbefore addition of a further portion of catalyst (5 g) and hydrogenatedas before for a further 2 hours after which no starting materialremained. The solution was filtered, evaporated to dryness under vacuumand the residue chromatographed on silica eluting with methylenechloride then methylene chloride:methanol 9:1. The crude product wasthen chromatographed again on silica (200 g) eluting with methylenechloride:methanol 19:1 to give after evaporation of the solvent undervacuum impure 22,23-dihydro-25-cyclohexylavermectin B1 as a brown foam(10 g). This material was dissolved in a mixture of isopropanol (200 ml)and sulphuric acid (2 ml ) and the brown solution was stirred at roomtemperature for 15 hours then poured into a mixture of ice and water(500 ml) and extracted with methylene chloride (3×200 ml ). The organiclayer was washed with saturated potassium hydrogen carbonate solution(100 ml), water (2×50 ml ) dried over anhydrous magnesium sulphate andevaporated under vacuum to give a crude gum which was chromatographed onsilica eluting with methylene chloride then methylene chloride:ethylacetate 2:1 to give 22,23-dihydro-25-cyclohexylavermectin B1monosaccharide. This compound was dissolved in anhydrous diethyl etherand the solution stirred with manganese dioxide to yield the titleproduct.

We claim:
 1. A compound of formula (I): ##STR6## wherein the broken linerepresents an optional bond, R¹ and R⁴ being absent when this bond ispresent, R¹ and R⁴ are independently H or OR¹⁴ where R⁴ is H, C₁ -C₈alkyl, C₂ -C₈ alkenyl, aralkyl, C₂ -C₈ alkanoyl, C₃ -C₈ alkenoyl,aralkanoyl, aroyl or carbamoyl;R² is:(a) an alpha-branched C₃ -C₈ alkyl,alkenyl, alkoxy-alkyl, or alkylthioalkyl group; an alpha-branched C₄ -C₈alkynyl group; a (C₅ -C₈)cycloalkyl-alkyl group wherein the alkyl groupis an alpha-branched C₂ -C₅ alkyl group; a C₃ -C₈ cycloalkyl or C₅ -C₈cycloalkenyl group, either of which is optionally substituted bymethylene or one or more C₁ -C₄ alkyl groups or halo atoms; or a 3 to 6membered oxygen or sulphur containing heterocyclic ring which issaturated, or fully or partially unsaturated and which is optionallysubstituted by one or more C₁ -C₄ alkyl groups or halo atoms; or (b) agroup of the formula --CH₂ R⁸ wherein R⁸ is H, C₁ -C₈ alkyl, C₂ -C₈alkenyl, C₂ -C₈ alkynyl, alkoxyalkyl or alkylthioalkyl containing from 1to 6 carbon atoms in each alkyl or alkoxy group, wherein any of saidalkyl, alkoxy, alkenyl or alkynyl groups are optionally substituted byone or more halo atoms; or R⁸ is a C₃ -C₈ cycloalkyl or C₅ -C₈cycloalkenyl group, either of which is optionally substituted bymethylene or one or more C₁ -C₄ alkyl groups or halo atoms; or R⁸ is a 3to 6 membered oxygen or sulphur containing heterocyclic ring which is besaturated, or fully or partially unsaturated and which is optionallysubstituted by one or more C₁ -C₄ alkyl groups or halo atoms; or R⁸ is agroup of the formula SR⁹ wherein R⁹ is C₁ -C₈ alkyl, C₂ -C₈ alkenyl, C₃-C₈ alkynyl, C₃ -C₈ cycloalkyl, C₅ -C₈ cycloalkenyl, phenyl orsubstituted phenyl wherein the substituent is C₁ -C₄ alkyl, C₁ -C₄alkoxy or halo; or R⁸ is a 3 to 6 membered oxygen or sulphur containingheterocyclic ring which is saturated, or fully or partially unsaturatedand which is optionally substituted by one or more C₁ -C₄ alkyl groupsor halo atoms; or (c) a C₁ -C₆ alkyl group substituted by one oxo or oneor more hydroxy groups or by a single oxygen atom on two adjacent carbonatoms forming an oxirane ring, or R² is a C₁ -C₅ alkyl group substitutedby a (C₁ -C₆) alkoxycarbonyl group, said substituents on R₂ beingattached to either or both of a terminal carbon atom and a carbon atomadjacent a terminal carbon atom of R² ; or (d) ═CH₂ or a group of theformula: ##STR7## wherein R¹⁰ and R¹¹ are both H; R¹⁰ is H and R¹¹ is C₁-C₃ alkyl, or one of R¹⁰ and R¹¹ is H and the other is phenyl,heteroaryl, C₂ -C₆ alkoxycarbonyl or substituted phenyl or heteroarylwherein said substituent is fluorine, chlorine, C₁ -C₄ alkyl, C₁ -C₄alkoxy, C₁ -C₄ alkylthio, hydroxy(C₁ -C₄)alkyl, cyano, aminosulphonyl,C₂ C₆ alkanoyl, C₂ -C₆ alkoxycarbonyl, nitro, trifluoromethyl,trifluoromethoxy, amino or mono or di(C₁ -C₄) alkylamino; and X is adirect bond or is an alkylene group having from 2 to 6 carbon atomswhich is straight or branched-chain; or (e) phenyl which may optionallybe substituted with at least one substituent selected from C₁ -C₄ alkyl,C₁ -C₄ alkylthio groups, halo atoms, trifluoromethyl, and cyano; or R²is a group of formula (II): ##STR8## wherein Z is O, S or --CH₂ -- anda, b, c and d are each independently 0, 1 or 2, the sum of a, b, c and dnot exceeding 5; R⁶ is H or C₁ -C₆ alkyl; R⁷ is CH₃, --CH₂ --OR¹⁴ whereR¹⁴ is as defined above, or --CH₂ X where X is a halogen atom; and R³ isa group of formula: ##STR9## wherein R⁵ is attached to C-4" or C-4' by asingle bond and is hydrogen, halo, hydroxy, C₂ -C₉ alkanoyloxy, C₃ -C₉alkenoyloxy, aroyloxy, C₁ -C₈ alkoxy, amino, N-(C₁ -C₈)alkylamino,N,N-di(C₁ -C₉)-alkylamino, N-(C₂ -C₉)alkanoylamino, or N,N-di(C₂-C₉)alkanoylamino; or R⁵ is attached to C-4" or C-4' by a double bondand is oxo, optionally substituted oximino, semicarbazido, N-(C₁-C₄)alkylsemicarbazono, N,N-di(C₁ -C₄)alkylsemicarbazono, (C₁-C₄)alkylbenzoylhydrazono, and R¹² and R¹³ are independently H, CN,CONH₂, C₁ -C₆ alkyl or aryl optionally substituted with at least onehalo, OH, C₁ -C₆ alkoxy or C₁ -C₆ alkylthio group.
 2. A compoundaccording to claim 1, in which R⁴ is H and R¹ is OH.
 3. A compoundaccording to claim 1 in which R² is isopropyl, sec-butyl or cyclohexyl.4. A compound of claim 1 in which R⁵ is --OH.
 5. A compound of claim 1in which one of R¹² and R¹³ is H and the other is H, methyl, cyano orcarbamoyl.
 6. A compound according to claim 1 whichis:5-cyanomethylidene-25-cyclohexyl avermectin B2;5-carbamoylmethylidene-25-cyclohexyl avermectin B2;5-cyanomethylidene-22,23-dihydroavermectin B1a monosaccharide;5-methylidene-22,23-dihydroavermectin B1a monosaccharide;5-methylidene-25-cyclohexyl-22,23-dihydroavermectin B1 monosaccharide;5-methylidene25-cyclohexylavermectin B2; or5-ethylidene-25-cyclohexylavermectin B2.
 7. A pharmaceutical compositioncomprising an effective amount of a compound of claim 1 and apharmaceutically acceptable carrier or diluent.
 8. A method of treatingparasitic infections in a mammal comprising administering to said mammalan antiparasitic effective amount of a compound of claim
 1. 9. A methodof preventing parasitic infections in a mammal comprising administeringto said mammal an antiparasitic effective amount of a compound ofclaim
 1. 10. A method of making a compound of formula (I) as defined inclaim 1 which comprises reacting a compound of formula (II), ##STR10##wherein R¹, R², R³, R⁴, R⁶ and R⁷ are as defined in claim 1 with acompound of formula R¹³ R¹² C═PPh₃ where R¹² and R¹³ are as defined inclaim 1 and Ph is phenyl.
 11. A method according to claim 10 in whichthe reaction takes place in an inert organic solvent at a temperaturefrom -100° C. to 0° C.
 12. A method of claim 11 in which R¹² or R¹³ iscyano and the cyano group is subsequently converted to a carbamoyl groupby hydrolysis.
 13. A method according to claim 10, in which R¹² or R¹³is cyano and the cyano group is subsequently converted to a carbamoylgroup by hydrolysis.